This application proposes a novel combination of magnetic resonance (MRS) and near infrared optical (NIRS) spectroscopy for studying energy metabolism in normal and abnormal human muscle. Our goal is to develop diagnostic procedures for human muscle function from principles of energy balances and their regulation developed by prior work on this grant. Metabolism, energetics and function of muscle are tightly integrated and provide crucial information needed to define normal and interpret abnormal muscle function in the intact human limb. In this application we add mouse muscle studies to test efficacy and accuracy of our quantitative spectroscopic methods. The first three specific aims exploit the notions of an integrative mass and energy balance developed in prior years of this grant to quantify the role of oxygen flux in the control of oxidative and glycolytic energy metabolism. First, we ask how mitochondria link O2 with ATP fluxes in oxidative phosphorylation (Aim 1). We evaluate the coupling between phosphorylation and oxidation with experiments that focus on cellular respiration in intact human muscle as a function of oxygen tension. These experiments are likely to establish new views of oxygen supply to mitochondria and the role of intracellular partial pressure of oxygen in the regulation of oxidative phosphorylation by [ADP]. Second, we examine how muscle diffusion conductance links O2 delivery with mitochondrial demand (Aim 2). This aim compares the magnitude of the oxygen pressure gradient versus the diffusive conductance in oxygen uptake flux. Third, we evaluate how much the diffusion conductance for oxygen uptake influences the balance between glycolytic flux and H+ & lactate efflux (Aim 3). If our hypothesis is supported, diffusive conductance will be found to play a similar functional role for H+ & lactate balances and for oxygen balance. Finally, Aim 4 shows how this integration uniquely accounts for abnormal muscle energetics in aging. Thus this project links all the major components of energy balance governing the supply of ATP to the demands of cellular ATPases into a coherent whole and tests how alteration of these linkages and their regulation influences function, using elderly muscle as an example.